Systems and methods for microscopic analysis of biological material have been used for characterization and diagnosis in many applications. Fluorescence microscopy, for example, has been used for optical analysis including the histological analysis of excised tissue specimens. Optical coherence tomography has been used for three dimensional imaging of tissue structures, however, the limited resolution of existing systems has constrained its use for definitive pathological analysis. Confocal microscopy has been used for high resolution imaging and has controllable depth of field but limited imaging speed.
Multiphoton microscopy is based on the nonlinear excitation of fluorophores in which fluorescence generation is localized at the focus of excitation light. Multiphoton microscopy is used for deep tissue imaging because of its subcellular three dimensional (3D) resolution, minimal phototoxicity, and tissue penetration depth of over a few hundred micrometers. It has become useful in biomedical studies such as neuronal plasticity, angiogenesis in solid tumors, transdermal drug delivery, and non-invasive optical biopsy, for example.
A practical limitation of multiphoton microscopy is its imaging speed which typically lies in a range of less than two frames per second. While this speed is sufficient in many cases, there remain applications in which can be enhanced by improvements in imaging speed. There is a continuing need for further improvements in microscopic analysis of biological materials for numerous applications.